The structure of the 21 S latent activity dynein-1 (LAD-1) particle has been investigated by limited proteolytic cleavage with trypsin and with chymotrypsin. The A alpha and A beta heavy polypeptide chains show different characteristic digestion patterns which remain essentially unchanged whether the chains are components of the 21 S LAD-1 particle or are in the form of separated fractions, although changes in their relative digestion rates upon separation suggest that the A beta chain in the 21 S particle is partially protected from digestion by the presence of the A alpha chain and intermediate chains 2 and 3. The progressive digestion of the A chains and intermediate chains causes an eventual dissociation of the 21 S particle to smaller particles sedimenting in the range 10 to 14 S. Within this broad peak, the fragments from the A alpha chain peak in the 10 to 12 S region, while those from the A beta chain peak in the 12 to 14 S region. Digestion of whole axonemes to a stage at which the A alpha chain is substantially digested but the A beta chain remains mostly intact, enables a large amount of 21 S dynein-1 to be solubilized by 3 mM MgATP2(-) in the presence of 0.1 M NaCl, pH 7.0. This indicates that the affinity of the 0.6 M NaCl-sensitive bond of the outer arm to the A-tubule is diminished substantially by the early stages of digestion of the A alpha chain.